Objective To screen the conservative,stable and specific DNA signature sequence in the plasmid of Yersinia pestis.Methods Specific validation trials and stability of the qualification test were carried out to 40 strains of Yersinia pestis,47 strains of non-Yersinia pestis of home and wild types of rodent in Yunnan,by using 32 DNA sequences derived from Yersinia pestis in the plasmid and conventional PCR technology,and Yersinia pestis vaccine strain EV76 as a positive control.Results Four pairs of relatively conservative,stable and specific DNA marker genes were screened:YPMT1.05c,YPMT1.03c,YPMT1.42 and YPMT1.04c.Conclusions The 4 pairs of Yersinia pestis DNA signature sequences can be used for rapid diagnosis of plague.

Objective To explore the feasibility of culturing dermal papillae cells (DPC) of hu- man hair in a serum-flee medium,and to observe the growth characteristics of these cells.Methods Cell culture flasks (plates) were pretreated with fibronectin,and DPC (2nd passage) were incubated with Williams E serum-flee medium supplemented with insulin-transferrin-selenite (ITS).Cells were observed by an inverted phase-contrast microscope.Proliferation of DPC was evaluated with 3-(4,5-dimethylthia- zol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay and by their growth curve.Results In a serum-free medium,2nd passage DPC adhered to the flask surface within two to four hours of incubation; two to three days later,confluence,of the cells was observed,without noticeable proliferation.Four days later,cell connection was interrupted,isolated cells or cell clusters were seen,and detachment of some cells from the flask surface was observed.One to two weeks later,most cells had died.After incubation with 4% bovine serum for ten hours,cell proliferation was observed surrounding the remaining viable cell colonies. DPC growth curve showed stagnant phase and slow growth phase;however,log growth phase was not ob- served.Conclusion DPC could be successfully cultured in serum-free medium.However,the culture con- dition needs to be further optimized.

Objective To compare the difference of biochemical characteristics and virulent Pst Ⅰ of Yersinia pestis strains in traditional focuses of plague in Yunna Province and in the new focuses of plague in Yulong County. Methods The identification data of biochemical characteristics(Rhamnose, Glycerol, Maltose, L-Arabina and Melibiose fermentation) and virulence factor(Pst Ⅰ) from different focuses of plague in Yunna Province were Retrospectively collected by tube test followed by the analysis using statistics software SAS 8.0 by Fisher exact probability of disordered two-way R × C table χ2 test. Results Among 48 strains of Yersinia pestis from hantaan type plague focus, 1 strain fermented L-maltose, 48 strains fermented Glycerol. Among 165 strains of Yersinia pestis from the Soul type plague focus, 1 strain did not ferment L-maltose, only one of them fermented Glycerol. 1 strain from the Soul type plague focus was confirmed to have mutation, for the test of nitrate reduction reaction was negative. All 5 strains of Yersinia pestis from the new focuses of plague in Yulong County fermented L-maltose and Glycerol. The statistical result showed that the differences in L-maltose and Glycerol fermentation of Yersinia pestis from different natural focuses of plague in Yunnan Province were statistically siguificant (P < 0.01). The differences of other biochemical characteristics and Pst Ⅰ were not statistically significant (P > 0.01). Conclusions Biochemical characteristics of Yersinia pestis from the hantaan type plague focus and the Soul type plague focus in Yunnan province are overlapping. Biochemical characteristics of Yersinia pestis from the new focuses of plague in Yulong County are different from those tradition focuses of plague in Yunna Province but share similarities to those from Unquiculatus focuses in North Tibet.

Antibodies, Monoclonal, Murine-Derived ; metabolism ; Diagnosis, Differential ; Female ; Follow-Up Studies ; Hemangioendothelioma ; metabolism ; pathology ; surgery ; Hemangioendothelioma, Epithelioid ; metabolism ; pathology ; surgery ; Humans ; Lymphatic Metastasis ; Middle Aged ; Platelet Endothelial Cell Adhesion Molecule-1 ; metabolism ; Sarcoma ; metabolism ; pathology ; surgery ; Thigh ; Vimentin ; metabolism ; von Willebrand Factor ; metabolism

Adult ; Antiphospholipid Syndrome ; diagnosis ; Cesarean Section ; Diagnostic Errors ; Female ; Humans ; Infarction ; diagnosis ; Liver Diseases ; diagnosis ; Pregnancy

Adolescent ; Adult ; Aged ; DNA, Circular ; blood ; Female ; Fluorescence ; Hepatitis B virus ; genetics ; isolation & purification ; Hepatitis B, Chronic ; virology ; Humans ; Male ; Middle Aged ; Polymerase Chain Reaction ; methods ; Sensitivity and Specificity

OBJECTIVETo investigate the expression and regulation mechanism of mismatch repair (MMR) genes in chronic myeloid leukemia (CML).

METHODSExpression of MMR genes hMSH2, hMSH3, hMSH6, hMLH1 and hPMS2 mRNAs in 62 CML patients and K562 cell line were detected by semi-quantitative reverse transcription polymerase chain reaction (RT-PCR). Expression of bcr-abl mRNA and MMR genes mRNA were detected by RT-PCR in 26 CML patients with allogeneic peripheral blood stem cell transplantation (allo-PBSCT) and 4 CML patients on imatinib treatment. Expression of bcr-abl mRNA was detected by RT-PCR and tyrosine phosphorylation of BCR-ABL fusion protein by Western blot.

RESULTSExpression of hMSH2, hMSH3 and hMLH1 mRNA was significantly lower in CML and K562 cells than in normal control (P < 0.05). In 26 CML with allo-PBSCT and 4 CML patients on imatinib treatment, expressions of hMSH2, hMSH3 and hMLH1 mRNA was enhanced while expression of bcr-abl mRNA decreased. In CML MNC after imatinib treatment and in K562 cells, expression of hMSH2, hMSH3 and hMLH1 mRNA was enhanced while tyrosine phosphorylation of BCR-ABL fusion protein decreased.

CONCLUSIONExpressions of hMSH2, hMSH3 and hMLH1 mRNA were down-regulated by bcr-abl fusion gene.

Adult ; Aged ; Antineoplastic Agents ; pharmacology ; Benzamides ; DNA Mismatch Repair ; Female ; Fusion Proteins, bcr-abl ; genetics ; metabolism ; Gene Expression ; Gene Expression Regulation, Neoplastic ; Humans ; Imatinib Mesylate ; K562 Cells ; Leukemia, Myelogenous, Chronic, BCR-ABL Positive ; genetics ; Male ; Middle Aged ; Piperazines ; pharmacology ; Pyrimidines ; pharmacology ; RNA, Messenger ; genetics ; Reverse Transcriptase Polymerase Chain Reaction

This study was aimed to investigate the expression and regulation mechanism of DNA-dependent protein kinase catalylic subunit (DNA-PKcs) in chronic myeloid leukemia (CML) and its role in blast crisis of CML. Expression of DNA-PKcs mRNA was detected by semi-quantitative reverse transcription polymerase chain reaction (RT-PCR) and DNA-PKcs protein by Western blot in 62 CML patients and K562, as compared to those of 23 normal individual controls. In 26 CML patients received allogeneic peripheral blood stem cell transplantation (allo-PBSCT) and 4 CML patients treated with imatinib, the expression of bcr-abl mRNA and DNA-PKcs protein was detected by RT-PCR and Western blot, respectively. After treatment with imatinib in mononuclear cell (MNC) of CML patients and K562 in vitro, expression of DNA-PKcs mRNA was detected by RT-PCR and DNA-PKcs protein level, tyrosine phosphorylation of bcr-abl fusion protein were detected by Western blot. The results showed that the expression of DNA-PKcs protein was significantly lower in CML and K562 than those in normal control (P<0.05). In 26 CML patients received allo-PBSCT and 4 CML patients treated with imatinib, the expression of DNA-PKcs protein was enhanced while the expression of bcr-abl mRNA decreased. After treatment of MNC of CML and K562 with imatinib in vitro, the expression of DNA-PKcs protein was enhanced while tyrosine phosphorylation of bcr-abl fusion protein decreased. It is concluded that the expression of DNA-PKcs protein is down-regulate by bcr-abl fusion gene, and the bcr-abl fusion gene down-regulate the expression of DNA-PKcs protein by post-transcriptional mechanism; the decrease of DNA-PKcs protein expression may be one of mechanisms underlying the acute transformation of CML.
Adult ; Aged ; Benzamides ; Bone Marrow Cells ; metabolism ; DNA-Activated Protein Kinase ; biosynthesis ; genetics ; Female ; Fusion Proteins, bcr-abl ; biosynthesis ; genetics ; Humans ; Imatinib Mesylate ; Leukemia, Myelogenous, Chronic, BCR-ABL Positive ; genetics ; therapy ; Male ; Middle Aged ; Peripheral Blood Stem Cell Transplantation ; Piperazines ; therapeutic use ; Pyrimidines ; therapeutic use ; RNA, Messenger ; biosynthesis ; genetics

OBJECTIVETo investigate the expression dynamics and significance of matrix metalloproteinase-2 (MMP-2) membrane type-matrix metalloproteinase-2 (MT-MMP-2) in hepatic fibrosis and its reversal counterpart.

METHODSAn experimental CCl4 induced hepatic fibrosis rat model was established by intraperitoneal administration of carbon tetrachloride for 2, 4, 6, 8, 10 weeks, and normal rats were used as a control group. The immunohistochemical methods and in situ hybridization were used to detect MMP-2,MT-MMP-2 mRNA and related antigens in the liver.

RESULTSMMP-2,MT-MMP-2 mRNA and related antigens were expressed in mesenchymal cells and parts of hepatocytes besides active pathological changes, especially in the fibrous septum and portal area. Expression of MMP-2,MT-MMP-2 mRNA and related antigens were increased in hepatic fibrosis and decreased gradually in its reversal counterpart.

CONCLUSIONThis study suggested that mesenchymal cells are the main cellular origins of MMPs. The levels of MMP-2 and MT-MMP-2 antigens and gene expression were closely related to hepatic fibrosis. MMP-2 and MT-MMP-2 may play important roles in hepatic fibrosis and its reversal counterpart.

Animals ; Carbon Tetrachloride Poisoning ; Gene Expression Regulation, Enzymologic ; Hepatocytes ; enzymology ; Liver ; enzymology ; pathology ; Liver Cirrhosis, Experimental ; enzymology ; etiology ; pathology ; Male ; Matrix Metalloproteinase 2 ; biosynthesis ; genetics ; Matrix Metalloproteinases ; biosynthesis ; genetics ; Matrix Metalloproteinases, Membrane-Associated ; Mesenchymal Stromal Cells ; enzymology ; RNA, Messenger ; biosynthesis ; genetics ; Rats ; Rats, Wistar

In this paper, we studied the relationship between the prostaglandin F(2alpha) (PGF(2alpha))-induced cardiac hypertrophy and calcineurin (CaN) signal transduction pathway in vivo and in vitro. Male Sprague-Dawley rats were given a single i.p. injection with monocrotaline (MCT) (60 mg/kg) and then given orally with celecoxib (20 mg/kg) or vehicle once a day for 14 d before (from d 1 to d 14) or after (from d 15 to d 28) right ventricular hypertrophy (RVH) was formed. Body weight (BW), right ventricular weight (RV), left ventricular with septum weight (LV), as well as lung weight were determined. RVH index (RVHI=RV/LV), RV/BW, and lung weight/BW were calculated and histological changes were observed with transmission electron microscope. PGF(2alpha) level, atrial natriuretic peptide (ANP) and CaN mRNA expressions, expression of CaN and its downstream effectors, NFAT(3) and GATA(4) protein were assayed by EIA kit, RT-PCR, and Western blotting, respectively. The cardiomyocyte hypertrophy in primary culture induced by PGF(2alpha) (0.1 micromol/L) was evaluated by measuring the cell diameter, protein content, and ANP mRNA as well as CaN mRNA expressions. It was found that 14 d or 28 d after MCT was given, the RVHI, RV/BW, and lung weight/BW were significantly increased by 47%, 53% and 118%, and by 64%, 94% and 156%, respectively; at the same time PGF(2alpha) levels in RV tissue were increased by 44% and by 51% with increasing RVHI, and elevated expressions of ANP and CaN mRNA, as well as CaN, NFAT(3) and GATA(4) proteins in a positive correlation manner. Furthermore, some histological injuries were found in RV tissue. Celecoxib, a cyclooxygenase inhibitor, obviously blunted the elevation of RVHI, RV/BW, and lung weight/BW no matter it was given before or after RVH. In vitro experiments showed that 0.1 micromol/L PGF(2alpha) significantly increased the cardiomyocyte diameter and protein content, and promoted ANP and CaN mRNA expressions, which was blocked by cyclosporin A, a CaN inhibitor. Our results indicate that PGF(2alpha) may be involved in cardiac hypertrophy induced by MCT in rats through CaN signal transduction pathway.
Animals ; Calcineurin ; genetics ; metabolism ; physiology ; Cells, Cultured ; Dinoprost ; metabolism ; physiology ; Hypertrophy, Right Ventricular ; chemically induced ; metabolism ; physiopathology ; Male ; Monocrotaline ; Myocytes, Cardiac ; metabolism ; pathology ; RNA, Messenger ; genetics ; metabolism ; Rats ; Rats, Sprague-Dawley ; Signal Transduction ; physiology